Neomycin-induced Autophagy In Prostate Cancer PC-3 Cells

Objective: This experiment aimed to observe if novobiocin inhibit the proliferation of PC-3 cells in vitro,to study the cell autophagy induced by novobiocin in the PC-3 prostate cancer cell line,to find the possible mechanisms and to provide experimental and theoretical basis for novobiocin treatment of CRPC.Methods: The PC-3 prostate cancer cell line was cultured with 0mmol/l、0.2mmol/l、0.4 mmol/l、0.8 mmol/l、1.2 mmol/l、1.6 mmol/l novobiocin in culture medium in vitro for 24 h,and was divided into treated groups and control group.The inhibitaed rate of cell proliferation of PC-3 cells was assayed by MTT among different novobiocin concentration groups.The ultra microstructure transformation of cells were observed by transmission electron microscope(TEM),different amounts of autophagosome were observed between the experimental group and the control group.The level of protein expression of LC3-Ⅱ 、Beclin1 and P62 were examined by immunofluorescence.The level of m RNA of autophagy gene LC3-Ⅱ and Beclin-1 was examined by RT-PCR among the experimental groups and the control group.The level of protein expression of LC3-Ⅱ、Beclin1 and P62 was examined by western blot.Results: 1.Within the concentration of(0-1.6)mmol/l,the proliferation rate of PC-3 in vitro was obviously inhibited by novobiocin.As thenovobiocin concentration increasing,the inhibition ratio was stepping up and the half inhibition ratio was 1.52mmol/L.2.The autophagosome with double-membrane structure can be examined by the TEM,and the number was stepping up as the concentration increasing.3.Immunofluorescence showed the expression of P62 was decreased in PC-3 cells after treated with novobiocin,and the expression of LC3-Ⅱ was increased.4.RT-PCR showed the level of m RNA of Beclin-1 and LC3-Ⅱ was increased by novobiocin with the increase of the concentration of novobiocin.5.Western blot showed the level of protein expression of LC3-Ⅱ and Beclin1 was increased,the level of protein expression of P62 was decreased.Conclusion: 1.We could find that novobiocin inhibit the proliferation of PC-3 cells in vitro by a dose-dependent manner.2.Within a certain drug concentration,the level of expression m RNA and protein of LC3-Ⅱ and Beclin1 in PC-3 cells could be improved by novobiocin,the level of protein of P62 was decreased.3.The proliferation rate of PC-3 cells could be inhibited by novobiocin inducing autophagy cell death.